Phosphate ester groups in proteoglycans from bovine nasal cartilage.

1. Proteoglycan subunits isolated by standard procedures from bovine nasal cartilage, previously incubated in the presence of [32P]phosphate contain [32]-phosphate ester groups as a regular structural component. 2. Contamination of the proteoglycan subunit with 32P-labeled nucleic acids could be excluded by repeated cesium chloride density gradient centrifugation under associative and dissociative conditions, lanthanum chloride precipitation, gel filtration and by the resistance of the proteoglycan subunit associated 32P to phosphoric diester hydrolases. 3. The [32P]phosphate ester groups are associated to the chondroitin sulfate peptide fraction obtained by proteolytic digestion of the proteoglycan subunit molecule. Degradation of the chondroitin sulfate peptide by chondroitinase ABC resulted in a 32P-labelled oligosaccharide peptide fraction, that contains xylose, galactose, glucuronic acid and inorganic phosphate in a molar ratio 1 : 2 : 1 : 0.12. 4. 32P radioactivity is released as inorganic phosphate by treatment of the 32P-labelled oligosaccharide peptide with acid phosphatase or alkali.

Histochemical properties of cartilage proteoglycans.

Proteoglycan interaction with alcian blue at different concentrations of magnesium chloride was studied both in vitro and in histological sections of paraffin-embedded tissues. Our experiments indicate that a) proteoglycans with different contents of chondroitin sulfate and keratan sulfate, prepared under nondegradative conditions, are not distinguishable on the basis of the critical electrolyte concentrations at which staining is abolished; b) the state of aggregation of proteoglycans only very slightly affects the alcian blue affinity of the macromolecules at different concentrations of magnesium chloride; c) the interaction of proteoglycans with other components of the connective tissue matrix is an important factor in determining the strength of binding of alcian blue to the polyanionic macromolecules in histological sections. These factors should be considered in interpreting histochemical data obtained by staining tissue sections with alcian blue at different concentrations of magnesium chloride. Proteoglycans, like glycosaminoglycans, are only weakly periodic acid-Schiff-positive.

Endonasal venular permeability in rats exposed to the cold.

Sixteen rats were killed 22 hrs after being injected with a dose of 4 mCi(198)Au mixed with 16 mg of inert colloidal gold/100g body wt. The anterior nasal mucosa of the septum and turbinates in 10 rats which were injected and kept in a temperature of 5-7 degrees C showed mean radioactive counts 2-3 times higher than those recorded in the same areas in 6 control rats kept in ordinary room temperature (22 degrees C). In all sixteen rats the highest counts were obtained in the RES organs, namely the liver, spleen, adrenals and the bone marrow, in descending order. Other tissues revealed no significant radioactivity except for minor counts in the lungs and kidneys. The importance of these findings regarding the localisation of granulomatous diseases and vasculitis in the nose is discussed.

The fate of homologous nasal septal cartilages in tympanoplasty.

We have used homologous nasal septal cartilage for tympanoplasty for the last 8 years and obtained satisfactory results. In order to demonstrate the fate of homograft cartilage implanted into the middle ear, mucopolysaccharides have been studied by means of enzyme digestion. The matrix of normal septal cartilage was divided into three regions: 1) pericellular region; chondroitin sulfate B, 2) distal interstitial region; hyaluronic acid and chondroitin sulfates, 3) peripheral interstitial region; collagen. In preserved cartilage, chondroitin sulfate B was lacked out, but hyaluronic acid and collagen remained intact though the amount of mucopolysaccharides diminished slightly when compared with normal septal cartilage. Homograft cartilages evidenced depletion of mucopolysaccharides. Homograft cartilages should be used for the purposes mentioned, though not as material for columella, nor for reconstruction of large bone defect.

Nasal glands in newborn infants.

The entire mucosa of the nasal septum from four newborn infants was removed, stained by the PAS-alcian blue whole-mount method, and the density of mucous glands was determined in 12 different localities. The interindividual median density was 30.7 glands/mm2 and the median count 13,500 glands. The glands were regularly distributed over the entire respiratory region, but with a significant decrease in density in the posterior half. It has been demonstrated that all glands are laid down before birth, that their density decreases with increasing age until it reaches 8.4 glandss/mm2 at an adult age, and that acute or recurrent acute catarrhal or inflammatory changes of the nose do not lead to the newformation of glands.

[Organization of the stroma in the bony nasal septum and its homologues in representative skulls]. / Organizacja zrebu kostnego przegrody nosa i jej homologów u przedstawicieli czaszkowców.

In literature the notion and systematization of osseous nasal septum complex in man and other mammals are not fully and precisely verified. The examinations of the processes of ossification in ++bony cranium and nasal septum were being made in birds, fishes, reptiles and mammals. The skulls were macerated and the specimens were stained by use of alizarin. The radio- and photography were used for documentation. The results proved the opinion that all the osseous elements of nasal septum in mammals derived from the interorbital septum and axially palatal complex in fishes, reptiles and birds.

[Concentration of trace elements in the inferior turbinates and in the nasal septum of the human]. / Konzentration der Spurenelemente in der unteren Nasenmuschel und im Septum des Menschen.

The trace elements in 47 inferior turbinates (concha nasalis inferior) and in the nasal septum were examined by means of synchrotron radiation, spectroscopy of characteristic x-rays, and "sub-micron elemental mapping with the Oxford scanning proton microprobe", and the results compared. Both first methods displayed an age-dependent reduction of Zn and an increase of Pb concentration. In some specimens the As concentration was very high. The disposition of all elements except Zn, As and Fe was uniform. Zn and As were concentrated on the surface of the septum and Fe in the region of vascular channels.

The distribution of different molecular species of collagen in fibrous, elastic and hyaline cartilages of the pig.

The distribution of type II collagen, considered to be characteristic of cartilaginous tissues, was determined in various specialized cartilages of the mature pig. The tissues examined were: (1) fibrocartilage of the semilunar meniscus of the knee; (2) elastic cartilage of the external ear; (3) hyaline cartilage of (a) the synovial joint (b) the thyroid plate of the larynx, and (c) the nasal septum. The predominant species of collagen in each tissue, whether type I or type II, was appraised semi-quantitatively by analysis of purified collagen solubilized by pepsin and of peptide fragments produced by cyanogen bromide. Cyanogen bromide-derived peptides were characterized by column chromatography on CM-cellulose and by electrophoresis in sodium dodecyl sulphate-polyacrylamide gels. The proportion of each type of collagen was determined precisely by isolating the homologous small peptides alpha1(II)CB6 [nomenclature of Miller (1973) Clin. Orthop. 92, 260-280], by column chromatography on phosphocellulose and determining their relative proportions by amino acid analysis. Thus collagen of the fibrocartilage of the meniscus proved to be all type I; type II was not detected. In contrast, collagen of elastic cartilage of the outer ear, after rigorous exclusion of perichondrium, was type II. Similarly, type II was the only collagen detected in all the mature hyalline cartilages examined.

Properties of fractionated chondroitin sulphate from ox nasal septa.

1. Chondroitin sulphate was isolated from bovine nasal septa by precipitation with cetylpyridinium chloride after digestion of the tissue with papain. 2. The material was divided into two portions, one of which was partially degraded with testicular hyaluronidase. 3. Untreated and hyaluronidase-digested material were fractionated into a total of eleven subfractions by gel chromatography on Sephadex G-200 and Sephadex G-100 respectively. 4. Chemical analyses indicated that the composition of all the fractions was similar to that of chondroitin sulphate. However, electrophoresis revealed a charge-inhomogeneity in the low-molecular-weight fractions obtained after hyaluronidase digestion. 5. The physicochemical properties of the subfractions were investigated by sedimentation-velocity, diffusion and sedimentation-equilibrium studies, osmometry, viscometry and gel chromatography. The individual fractions were essentially monodisperse and showed molecular weights ranging from 2400 to 36000. 6. The relationship between the intrinsic viscosity and the molecular weight was [eta]=5.0x10(-6)xM(1.14), indicating that the chondroitin sulphate molecules assume a shape intermediate between that of a random coil and a stiff rod. 7. The relationship between the sedimentation constant and the molecular weight (>10(4)) was s(0) (20,w)=2.3x10(-2)xM(0.44).

Selective aggregation of proteoglycans with hyaluronic acid.

Conditions were established to separate proteoglycan aggregate (AH1) from a bovine nasal septum extract by associative rate zonal sedimentation on a NaCl gradient. The AH1 has a higher protein content than the mixed aggregate-monomer (A1) isolated by conventional associative CsCl density gradient centrifugation from a portion of the same extract. The same associative rate zonal conditions separated the A1 fraction into aggregated AH1 containing hyaluronic acid and nonaggregated proteoglycan monomer (N1) essentially free of hyaluronic acid. The AH1 fraction is richer in protein and keratin sulfate than is N1. Dissociative rate zonal sedimentation of A1 under conditions which totally sedimented most of the disaggregated monomer (AH1-D1) and the nonaggregated monomer N1 separated a less sedimentable protein and keratan sulfate-rich proteoglycan monomer (AH1-D2). Chromatography on Sepharose 2B under dissociative conditions demonstrated that the nonaggregated N1 monomer is intermediate in size between the disaggregated monomers AH1-D1 and AH1-D2. N1 has a buoyant density higher than AH1 and is practically equivalent to AH1-D1. All are dense fractions so that separation by CsCl density gradient equilibration is not feasible.

Investigation of molecular motion of proteoglycans in cartilage by 13C magnetic resonance.

13C nmr spectral parameters were measured for intact bovine nasal cartilage tissue, the purified proteoglycan aggregate, and chondroitin 4-sulfate. A comparison of integrated intensities obtained for four different samples of fresh tissue with an ethylene glycol standard indicated that at least 80% of the total glycosaminoglycan carbons in the tissue contributed to the spectrum. This result was confirmed by intensity measurements obtained at 56 degrees on fresh tissue and at 37 degrees after extensive papain digestion of fresh tissue. Spin lattice relaxation times and nuclear Overhauser enhancements were analyzed in terms of the following models of molecular motion: (a) single correlation time; (b) log X2 distribution of correlation times; and (c) anisotropic motion. The analysis indicates that the segmental motions of glycosaminoglycan chains are characterized by a broad distribution of correlation times centered at about 50 ns. Slow motion contributions to glycosaminoglycan line widths were reduced by dipolar decoupling (gammaH2/2pi = 65 kHz). Collagen intensity was observed in dipolar decoupled spectra, but not in scalar decoupled spectra of intact tissue, showing that the type II collagen in cartilage undergoes anisotropic motion like the type I collagen in tendon. Only glycosaminoglycan resonances were observed in spectra of a solution of proteoglycan aggregate before and after chondroitinase digestion. After subsequent digestion with papain, protein resonances were observed. These results suggest that the protein portions of the proteoglycan aggregate structure, in contrast with the glycosaminoglycan chains, have restricted backbone mobility and consequently a defined backbone structure.

The protein-polysaccharide complex of bovine nasal cartilage. Studies on the protein core.

1. Chondromucoprotein from bovine nasal cartilage was purified by cetylpyridinium chloride or by bismuth nitrate in acetone. 2. Amino acid compositions of crude and purified preparations were compared and few differences were found, in spite of the decrease in protein content on purification. 3. Amino acid analysis of bismuth-purified material revealed the existence of four groups of amino acids. Within each group, the amino acids were present in approximately equimolar concentrations. 4. Amino end-group assay on the same material showed six alpha-DNP derivatives. 5. A molecular weight of 6.3x10(5) for the protein-polysaccharide complex was calculated from the latter analysis.